Mitochondrial Calcium Uniporter Drives Chemoresistance in Pancreatic Cancer via Glutathione‐Mediated Stemness Maintenance

ABSTRACT Pancreatic ductal adenocarcinoma (PDAC) remains a lethal malignancy with poor prognosis due to chemoresistance. Using integrative single‐cell RNA sequencing, we identified that the upregulation of mitochondrial calcium uniporter (MCU) may contribute to chemoresistance and stemness maintenance in PDAC. MCU was highly expressed in chemotherapy‐resistant PDAC tumors and correlated with enhanced cancer stem cell properties. Mechanistically, MCU‐mediated mitochondrial Ca 2+ influx triggered endoplasmic reticulum (ER) stress and the downstream PERK‐eIF2α pathway. This cascade activated ATF4 and NRF2, which enhanced the transcriptional regulation of PSAT1 and SLC7A11. These changes promoted de novo glutathione (GSH) synthesis to scavenge reactive oxygen species (ROS) and sustain stemness. Genetic knockdown or pharmacological inhibition of MCU disrupted GSH synthesis, suppressed stemness, and restored sensitivity to nab‐paclitaxel plus gemcitabine (AG). High‐throughput screening identified MCU inhibitor NB‐598, which synergized with AG to inhibit tumor growth in preclinical models. These findings offer a potential novel therapeutic strategy to address chemoresistance in PDAC.